Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Aberrant Regulation of HDAC2 Mediates Proliferation of Hepatocellular Carcinoma Cells by Deregulating Expression of G1/S Cell Cycle Proteins

ABSTRACT: To investigate the specific roles of HDAC2 in the development of liver cancer, we employed large-scale gene expression analysis to identify the molecular signature that may affect enabling characteristics of cancer cells. Differentially expressed genes were analyzed on the Hep3B cells transfected with HDAC2 shRNAs, and recapitulated molecular signatures that related to hallmarks of cancer. With the aim of identifying changes in gene expression following gene knockdown of HDAC2, we performed expression profiling in shRNA expression vector-transfected Hep3B cells, and compared the results to those obtained from vector-transfected Hep3B cells.

ORGANISM(S): Homo sapiens

SUBMITTER: JUNGWOO EUN

PROVIDER: E-GEOD-32070 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:To investigate the specific roles of SIRT7 in the development of HCC, we employed large-scale gene expression analysis to identify the molecular signature that may affect enabling characteristics of cancer cells. Differentially expressed genes were analyzed on the Hep3B cells transfected with SIRT7 shRNAs, and recapitulated molecular signatures that related to hallmarks of cancer. For each of the experimental conditions, total RNA was extracted from three independent sets of the corresponding cell lines using TRIzol Reagent (Invitrogen), followed by clean up on Ambion columns (Illumina Total-Prep RNA Amplification Kit, Ambion). For each experimental condition, an RNA pool was then obtained by mixing equal quantities of total RNA from each of the three independent RNA extractions. Biotin-labelled cRNA targets were synthesized starting from 1.5 µg of total RNA. Double stranded cDNA synthesis was performed with Illumina® TotalPrep RNA Amplification Kit (Ambion), and biotin-UTP-labelled antisense RNA was transcribed in vitro using Ambion’s Kit. All steps of the labelling protocol were performed as suggested by Ambion (http://www.ambion.com/techlib/prot/ fm_IL1791.pdf). The size and the accuracy of quantitation of targets were checked by Experion (Bio-rad Laboratories., Hercules, CA) electrophoresis system, prior to and after cRNA purification. After purification, targets were diluted in hybridization buffer at a concentration of 240 ng/µl, and hybridization was allowed to proceed at 58°C for 20 h. For microarray analysis, the Illumina HumanHT-12 v4 Sentrix Expression BeadChip was used (Illumina, San Diego, CA). Hybridization of labelled cRNA to the BeadChip, washing, and scanning were performed according to the Illumina BeadStation 500× manual. The array signal was developed via 10-min incubation with streptavidin-Cy3. The HumanHT-12 v4 Sentrix Expression BeadChip was washed and subsequently dried via centrifugation for 4 min at a setting of 275 xg. The arrays were scanned on the Illumina BeadArray reader, a confocal-type imaging system with 532 (Cy3) nm laser illumination. Data from each sample was extracted with Genome Studio software (Illumina) using default parameters and then analyzed using GenPlex 3.0.

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Aberrant Regulation of HDAC2 Mediates Proliferation of Hepatocellular Carcinoma Cells by Deregulating Expression of G1/S Cell Cycle Proteins

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